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Staphylococcus aureus α-toxin modulates skin host response to viral infection  Lianghua Bin, PhD, Byung Eui Kim, MD, Anne Brauweiler, PhD, Elena Goleva,

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Presentation on theme: "Staphylococcus aureus α-toxin modulates skin host response to viral infection  Lianghua Bin, PhD, Byung Eui Kim, MD, Anne Brauweiler, PhD, Elena Goleva,"— Presentation transcript:

1 Staphylococcus aureus α-toxin modulates skin host response to viral infection 
Lianghua Bin, PhD, Byung Eui Kim, MD, Anne Brauweiler, PhD, Elena Goleva, PhD, Joanne Streib, BS, Yinduo Ji, PhD, Patrick M. Schlievert, PhD, Donald Y.M. Leung, MD, PhD  Journal of Allergy and Clinical Immunology  Volume 130, Issue 3, Pages e2 (September 2012) DOI: /j.jaci Copyright © 2012 American Academy of Allergy, Asthma & Immunology Terms and Conditions

2 Fig 1 Staphylococcal α-toxin, but not superantigens, enhances viral load in NHKs. NHKs were pretreated with indicated S aureus superantigens and α-toxin, followed by VV (MOI, 0.1) and HSV-1 (MOI, 0.1) inoculations. A and B, The viral mRNA expression levels of HSV-1 (Fig 1, A) and VV (Fig 1, B) were evaluated by using real-time PCR. C, Viral plaque formation was determined by using the viral plaque assay. The left panel shows crystal violet staining of viral plaques; the right panel shows quantitative results of viral plaques. All data are presented as mean ± SEM values. Data from 1 representative experiment of 3 independent experiments performed are shown. **P < .01 and ***P < .001. Journal of Allergy and Clinical Immunology  , e2DOI: ( /j.jaci ) Copyright © 2012 American Academy of Allergy, Asthma & Immunology Terms and Conditions

3 Fig 2 Sublytic α-toxin is the major staphylococcal virulent factor enhancing viral load in keratinocytes. A, Western blot showing α-toxin and TSST-1 protein expression in Hla-WT and Hla-KO S aureus culture supernatants. B, Viral gene expression in NHKs pretreated with supernatants collected from sham, Hla-WT, and Hla-KO S aureus cultures. Data are presented as mean ± SEM values. Data from 1 representative experiment of 3 independent experiments performed are shown. **P < .01 and ***P < .001. Journal of Allergy and Clinical Immunology  , e2DOI: ( /j.jaci ) Copyright © 2012 American Academy of Allergy, Asthma & Immunology Terms and Conditions

4 Fig 3 VV load is greater in murine skin inoculated with Hla-WT S aureus strain than with Hla-KO S aureus strain. A, Representative picture of murine skin lesions at day 7 of VV inoculation. B, Quantitative results of the sizes of murine skin lesions at day 7 of VV inoculation. Data are presented as mean ± SEM values. C, Representative images of VV immunofluorescent staining of murine skin lesion biopsy specimens taken at day 7 after VV inoculation (magnification ×40). D, VV immunofluorescent intensity. Data are expressed as mean ± SEM values. MFI, Mean fluorescence intensity. Journal of Allergy and Clinical Immunology  , e2DOI: ( /j.jaci ) Copyright © 2012 American Academy of Allergy, Asthma & Immunology Terms and Conditions

5 Fig 4 Silencing ADAM10 expression blocks sublytic α-toxin's effect on viral enhancement. A, Western blot showing ADAM10 protein level and α-toxin in indicated conditions. β-actin was used as a loading control. B, HSV-1 mRNA expression in indicated conditions. C, VV mRNA expression in indicated conditions. Data are presented as mean ± SEM values. Data from 1 representative experiment of 3 independent experiments performed are shown. Journal of Allergy and Clinical Immunology  , e2DOI: ( /j.jaci ) Copyright © 2012 American Academy of Allergy, Asthma & Immunology Terms and Conditions

6 Fig 5 H35A mutated α-toxin has no viral enhancement effect. NHKs were treated with indicated concentrations of wild-type (WT) α-toxin and H35A mutated α-toxin for 20 hours followed by 24 hours of viral incubation. Cells were then harvested for Western blot assay and real-time PCR. A, Western blot showing α-toxin in indicated conditions. β-actin was used as a loading control. B and C, HSV-1 (Fig 5, B) and VV (Fig 5, C) mRNA expression in indicated conditions. Data are presented as mean ± SEM values. Data from 1 representative experiment of 3 independent experiments performed are shown. Journal of Allergy and Clinical Immunology  , e2DOI: ( /j.jaci ) Copyright © 2012 American Academy of Allergy, Asthma & Immunology Terms and Conditions

7 Fig 6 α-Toxin promotes viral entry into keratinocytes. A, VV luciferase activity was measured in α-toxin–pretreated NHKs after incubation with VV luciferase at an MOI of 10 for 1 hour. B, NHKs were pretreated with or without α-toxin (20 ng/mL) for 20 hours, and cells were then subjected to 100 μmol/L acyclovir treatment for 2 hours. Finally, HSV-1 (MOI, 0.05) was added into the cells for additional 5- and 10-hour incubations. Intracellular HSV-1 DNA copies were evaluated by using real-time PCR. Data are presented as mean ± SEM values. Data from 1 representative experiment of 3 independent experiments performed are shown. Journal of Allergy and Clinical Immunology  , e2DOI: ( /j.jaci ) Copyright © 2012 American Academy of Allergy, Asthma & Immunology Terms and Conditions

8 Fig E1 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS) assay to analyze NHK survival in the presence of various concentrations of α-toxin. Journal of Allergy and Clinical Immunology  , e2DOI: ( /j.jaci ) Copyright © 2012 American Academy of Allergy, Asthma & Immunology Terms and Conditions

9 Fig E2 Expression of antimicrobial peptides, antiviral genes, and the HSV-1 receptor herpesvirus entry mediator C (HVEC) in NHKs after treatment with α-toxin was evaluated by using real-time PCR. Journal of Allergy and Clinical Immunology  , e2DOI: ( /j.jaci ) Copyright © 2012 American Academy of Allergy, Asthma & Immunology Terms and Conditions

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